Electrophoresis device

ABSTRACT

An electrophoresis device having a separating chamber, and two buffer containers one of which is attached to one end of said separating chamber and the other one is connected to the other end, and an electrode in each in said buffer containers, said device essentially having a box-shaped housing and a box area which is divided up into two partial areas serving as buffer containers by means of an essentially horizontal intermediate bottom of lesser depth than the box together with an intermediate wall which extends essentially vertically from the intermediate bottom.

United States Patent 1 1 Nees et al. Nov. 20, 1973 [5 ELECTROPiiORESISDEVICE 3,208,929 9/1965 Raymond et a1 204 299 3,374,166 3/1968 Ra 0nd...204 299 [751 Invenmrs f Hubert 3,563,880 2/197'1 An e i'son... 2041299Wilhelm Schmidt both of Erlangen, 3,579,433 5 1971 Dahlgren 204 299Germany [73] Assignee: Colora Messtechnik GmbH, Primary Examiner |ohn HMack Lorch/wuntw Germany Assistant ExaminerA. C. Prescott [22] Filed; No12, 7 Attorney--John Lezdey et al.

Appl. No.: 198,130

[30] Foreign Application Priority Data Nov. 14, 1970 Germany P 20 56129.9 Nov. 14, 1970 Germany P 20 56 128.8 Nov. 14, 1970 Germany P 20 56127.7

[52] US. Cl. 204/299, 204/180 G [51] Int. Cl B0 lk 5/00 [58] Field ofSearch 204/299, 180 G, 180 R, 204/301 [56] References Cited UNITEDSTATES PATENTS 2,853,448 9/1958 l-leiskell, Jr. 204/299 ABSTRACT Anelectrophoresis device having a separating chamber, and two buffercontainers one of which is attached, to one end of said separatingchamber and the other one is connected to the other end, and anelectrode in each in said buffer containers, said device essentiallyhaving a box-shaped housing and a box area which is divided up into twopartial areas serving as buffer containers by means of an essentiallyhorizontal intermediate bottom of lesser depth than the box togetherwith an intermediate wall which extends essentially vertically from theintermediate bottom.

36 Claims, 9 Drawing Figures PATENTED HUV 20 I375 SHIET l E? BACKGROUNDOF THE INVENTION This invention relates to an electrophoresis devicewith two buffer working tanks one of which is connected to one end of aseparating chamber and the other one of which is connected to the otherend of a separating chamber, and with one electrode each in the buffertanks.

Prior art electrophoresis devices are built in such a way that avertical separating chamber follows a first box-shaped container for thebuffer solution. A second buffer container, which can also bebox-shaped, is located on the vertical separating chamber. Theproduction of these components, which are preferably circular in shape,is expensive. Furthermore, because of the upper buffer tank, the overalldesign of the device os very unstable so that the danger of damage isquite great. In these prior art devices good accessibility to the upperand lower chamber limitation, which is necessary in variouselectrophoresis methods, is not present.

Also prior art preparative electrophoresis devices are essentiallyconstructed of axisymmetric parts they consequently have a cylindricallyshaped separating chamber, bowl-shaped buffer containers and an elutionchamber which is also circularly shaped in the cross section. The partsof the prior art devices are balanced to one another in such a way thatpreparative electrophoreses can be carried out. For the carrying out ofanalytical tasks, however, these devices are not suitable.

The yields of known preparative electrophoresis devices are alsorelatively low even in more modern devices. Also the selectivity. whichis achieved with known devices is still not optimum so that substanceswith practically the same electrophoretic rate of travel cannot besecurely separated.

OBJECTS OF THE INVENTION One object of this invention to create anelectrophoresis device in which these prior art disadvantages are notpresent, in which therefore a compact form of construction is available,simple parts can be used, the danger of damage from a low degree ofstability and difficult construction is excluded and a goodaccessibility of the upper and lower separating chambers is providedfor. It is a further object of this invention to provide a simple andefficient cooling of the separating chambers, as well as a hydrostaticpressure equalization in the two buffer containers without specialmeans, such as lifting pipes or similar, which are required for thispurpose.

It is a still further object of this invention to eliminate thedisadvantages of known preparative electrophoresis devices so that adevice is created with which higher yields and greater selectivities canbe achieved.

A still further object of the invention is to provide a device whichmakes it possible, without problems, to carry out not only preparativeelectrophoreses but also analytical electrophoreses.

A yet still further object of this invention is to permit an operationaccording to the most varied known elec trophoresis methods so that therange of application of the proposed device is expanded.

SUMMARY OF THE INVENTION The objects of this invention are achieved bymeans of an essentially box-shaped design of the electrophoresis device.The device of this invention is provided with a box-shaped area by meansof an essentially horizontal intermediate bottom which is of smallerdepth than the box, can be split up together with an intermediate wallwhich goes out from the intermediate bottom and extends essentiallyvertically to it into two partial areas serving as a buffer container.It is particularly advantageous to have the intermediate wall extendingvertically upwards into the box because when both partial areas are openupwardly ithe buffer fluids can be brought practically to the same levelwhereby the hydrostatic pressure equalization is brought about in thesimplest manner. The design according to this invention permits anarrangement of the separating chamber essentially in the plane of theintermediate wall. The thickness of the separating chamber may be variedin a particularly simple manner when parallel to the intermediate walland inthe region of the greater angle formed by the intermediate bottomand the intermediate wall. A plate can be applied which can beadjustable in its distance and which, together with the intermediatewall, forms the separating chamber.

The accessibility to the buffer containers and the separating chamber isparticularly simplified in that the front side wall of the box-shapedhousing and the lid of the housing'are removable. According to a furtherembodiment of this invention the device can be used for the applicationof the most varied electrophoresis methods, for example the continuouselectrophoresis method, the discontinuous electrophoresis method, theisoelectric focusing and the iso-tachoelectrophoresis when means forpressing in inserts (such as elution insert, isoelectric focusinginsert, and the like) in the partial space located below the separatingchamber to the lower limitation of the separating chamber. Thesepressing means are achieved preferably by means of a pressing devicewhich operates in the manner of a lifting stand.

This invention has a further advantage by permitting the cooling meanswhich are provided for cooling the separating chamber to engage thebuffer solutions in both buffer containers. It then suffices for thebuffer solutions to be cooled in the region of-the separating chambersince the buffer solutions themselves can act as a cooling medium.Within the framework of the invention it is advantageous to arrange alaminar cooling plate or hose parallel to the separating'chamber wallsin order to be able to achieve in this way the greatest possible coolingarea.

For this purpose the invention is based on the fact that with the knownconstruction principle of preparative electrophoresis devices neitherthe selectivity nor the yields can be increased to any considerabledegree. This is due to the fact that in the amplification of theseparating chamber diameter for increasing the yields the cooling of thecarrier substance becomes problematical and also the elution of greatercircular-shaped or circular-ring shaped cross sections can no longeroccur in a satisfactory manner. Inadequate cooling of the carriersubstance in the separating chamber results namely in the fact that atemperature gradient occurs between the inside, the center of thecylindrical separating chamber and its cooled outside wall.

Since the rate of travel in the separating chamber depends upon thetemperature and the flushing occurs with different flow velocities it isnot possible to increase the quality of the electrophoresis devices whenmaintaining the known principal design.

According to this invention the separating chamber has an essentialrectangular cross section and the elution chamber is attachable to oneend of the separating chamber. By means of the steps according to thisinvention it becomes possible to cool large separating chamber volumesefi'ectively owing to their essentially greater surfaces and also tocarry out the flushing on the elution side of the separating chamberwith uniform flow velocities. With this, however, the preconditions fora considerable improvement in quality of the known preparativeelectrophoresis devices are given. The rectangular separating chamberconstruction can be considerably simplified and made more economical incomparison to most of the blown cylindrical separating chambers by meansof the utilization of simple glass plates. The rectangular separatingchamber design also permits a compact design of the entire deviceincluding the buffer container. In addition, by removing the elutionchamber the device can easily be used immediately for analyticalelectrophoresis. An interchanging of the elution chamber against otherattachments to be placed on the underside of the separating chambermakes it possible to operate according to various electrophoresismethods and simplifies the work in that, for example, by means of anattachment formed as a fastener, the separating chamber can be filledwith carrier substance without having to be removed from the device andthe electrophoresis can immediately be started in the device after thepolymerization and the removal of the fastener. attachment without thedisturbing transport of the carrier substance and without anydifficulties of installation into the device.

Within the framework of this invention it is appropriate to constructthe elution chamber in such a way that its form essentially correspondsto the lower separating chamber opening, i.e., so that the elutionchamber is rectangular. lt is advantageous to construct the elutionchamber out of a plate which has a recess which corresponds to the lowerseparating chamber opening so that the lower separating chamberlimitation should be tightly attachable to the upper side of the plateover the recess. When the recess corresponds to the diameter of theplate, the plate is slit, and means are provided for closing the lowerside of the slit opposite the separating chamber. Connections for thefeeding and removal of the elution fluid are provided in the region ofthe small sides of the recess or of the slit. This procedure permitsuniform flow velocity over the entire cross section of the chamber,therefore uniform washing out of the electrophoretically separatedsubstances and consequently a high degree of selectivity on the part ofthe device. The feeding and removal of the elution fluid on the narrowsides of the rectangular elution chamber also causes a highconcentration of the eluted substance.

The slit may be provided with a semipermeable membrane as a means forclosing the separating chamber of the plate. This membrane is preferablysqueezed, by means of a second plate, between the second plate and thefirst plate. In this manner the second plate can have a correspondingrecess as in the first plate. Preferably, in the region of the narrowsides of the recess of the second plate, attachments are provided forthe feeding and removal of a buffer fluid hwich is particularly highlyconcentrated within the framework of the invention. The recess inthesecond plate can also be formed as a slit, whereby the slit on theside opposite that of the elution chamber is closed by means of a secondmembrane. In a similar manner the second membrane is also squeezed, bymeans of a third plate which is also slit, between that plate and thesecond plate. There is thus obtained a plate stack composed of threeplates with a slit which passes through and which corresponds in size tothe separating chamber outlet, whereby a continuous semipermeablemembrane is located betwen the plates. Such a plate stack is produced inthe simplest manner and combined with the elution chamber according tothe invention and also inserted into the device. For this purpose,within the framework of the invention, means are provided for thecompact pressing together of the stack and for the tight pressing of thestack against the bottom side of the separating chamher.

In the framework of the device, the elution chamber and the means forbearing pressure upon the elution chamber are preferably provided for inthe lower located region serving as the buffer container in the case ofa compact box-shaped construction of the electrophoresis devices inwhich, by means of an intermediate bottom and an intermediate wall, thehousing is divided up into two areas forming the buffer containerbetween which regions the separating chamber is located. The means forpressing against the elution chamber has a lifting stand which ispreferably formed of threaded bolts. These bolts are located on thebottom of the lower-situated region, lie vertically and the stand platecan move to and fro on these bolts by means of threaded nuts. In thismanner, the elution chamber can be pressed against the lower side of theseparating chamher.

It is particularly advantageous, for the cooling of the separatingchamber which is rectangular in cross section, to arrange the coolingagents parallel to the large sides of the separating chamber which canbe composed, for example, of a cooling house proceeding in a plane withimmersion frames.

In the case of the prior art electrophoresis devices, in the upper andlower buffer container, locking elements are provided for a frame inwhich an electrode is housed. These frames are placed immediately aboveand below the separating chamber. The placement below the separatingchamber has the disadvantage that gas bubbles rising up on the electrodereach the lower side of the separating chamber and in this way cause adisruption in the electrophoresis. in order to eliminate thisdisadvantage it is necessary to provide, between the lower side of theseparating chamber and the electrode in the buffer container, meanswhich prevent the gas bubbles from reaching the lower side of theseparating chamber. This is achieved by holding in another frame adeflecting means such as an absorptive paper which is shaped into anacute-angled groove in such a manner that the point points downwards andthe upper edges of the groove lie laterally outside of the lower limitof the separating chamber. Gas bubbles ris-,

ing up are pushed laterally upwards from the lower side of the grooveand cannot reach the separating chamher.

It can be seen that in this kind of construction the structure andassembly of the device are complicated and that there is the danger thatthe absorptive paper is mistakenly not inserted and that impurities canreach the buffer solution through the paper.

In accordance with a further embodiment of this invention at least thebuffer container, which is in contact with the lower end of theseparating chamber, has a recess in the inner container wall, preferablythe container backwall, for housing the electrode.

This feature prevents the rising gas bubbles from reaching the bottomside of the separating chamber. There is furthermore achieved that nospecial frame and no special holding means in the buffer container areneeded for the electrode so that the construction and erection of thispart of the electrophoresis device are considerably simplified.

Prior art devices also have a considerable disadvantage in that theseparating chamber is not easily accessible from above when the upperelectrode attachment is placed in the buffer container. The subsequentinsertion of the electrode attachment into the buffer container isdisadvantageous, however, since the buffer fluid is placed into motionthrough the insertion and thus the fluid substances which are piled upon the uppper side of the separating chamber can easily be mixed.Bearing this situation in mind, this invention makes it possible for theaccessibility to the upper limit of the separating chamber to beunhampered and also makes it possible to do away with a subsequentinsertion of the electrodes into the buffer 0containers.

This is achieved by also providing a recess for the electrode in thebuffer container which comes into contact with the upper end of theseparating chamber. This thereby provides the advantage of goodaccessibility and the elimination of the necessity of having to insertthe upper electrode subsequently.

Examples of construction of the invention are described in what followsbased on the drawing.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 shows a side view of theelectrophoresis device of this invention in which the side wall'has beenremoved: i g

FIG. 2 shows an alternate form ofa separating cham ber wall for use withthis invention;,

FIG. 3 shows the assembly of a separating chamber for use with thisinvention;

FIG. 4 shows a special design of a separating chamber part for use inthis invention;

FIG. 5 shows the elution attachment of the device of this invention;

FIG. 6 shows a special form of an elution chamber part for use with thisinvention;

FIG. 7 is a view of .the inside of the backwall of the electrophoresisdeviceof this invention;

FIG. 8 is a cross section view through the backwall of the deviceaccording to FIG. 7, and

FIG. 9 is a cross section view through an addition for working accordingto the method of iso-electric focus- DESCRIPTION OF THE PREFERREDEMBODIMENTS In FIG. 1 there is shown an electrophoreses device having abottom 1, lid 2, backwall3 and front side wall 4 of a box-shaped housing5 with a side-wall removed. Inside of the housing 5 an intermediatebottom 6 has been inserted into about one-third of the overall height ofthe housing. It extends beyond half of the depth of the fronting to thefront side wall 4. An intermediate wall 8 extends vertically upwardsfrom its free end 7. The intermediate bottom 6 and the intermediate wall8 provide the boundary, together with the backwall 3 and the side wall,not shown, of the housing 5, for a buffer container 9, and bound ittightly against the remaining area which forms a second buffer container10. Parallel to the intermediate wall 8 in the direction of the frontside wall 4 of the housing 5 with a distance, there is provided a plate1 1 which forms the separating chamber 12 of the electrophoresis devicetogether with the intermediate wall 8. The plate 1'] is somewhat higherthan the intermediate wall 8 so that the hatchedline-indicated buffersolution in the buffer container 9 can reach the upper end of theseparating chamber. The lower end of the separating chamber 12 issurrounded by the fluid which is located in the buffer container 10. Anelectrode 13 or 14 is provided in the backwall 3 in the region of thetwo buffer containers 9 and 10.

By means of the described construction of the device, in which accordingto the invention the two buffercontainers 9 and 10 are locatedessentially laterally, preferably on both sides of the separatingchamber 12, a pressed construction form is given, only simple parts areused, and there is no danger that the device will be exposed to the riskof damage because of inadequate stability or complicated construction.When the lid 2 is removed, the device is easily accessible from above sothat the extra shifting or layering of substances, which is often to beundertaken with great sensitivity, is easily possible. Also theintroduction of thebuffer solution from above into both buffercontainers is easily possible. Complicated and expensive pressureequalization devices are therefore not necewsary.

The front side wall 4 is detachable from the two side walls. thedetachability of the front side wall provides good access to the lowerend of the separating chamber 12 so that it can be filled with carriersubstance without having to be removed from the device, when the easilyaccessibly lower separating chamber opening is closed. The preparationof the carrier'substance can therefore take place inside of the device,whereas in the case of preivously known devices this had to be doneoutside of the device. The loaded separatingchamber can also remain inthe device during the time of the polymeriza tion of the carriersubstance.

The described construction also makes it possible to use the bufferfluid in the containers 9 and 10 as a cooling agent, in which a coolingdevice, for example cooling plates or cooling hoses l5, 16 are placedparallel to the surfaces 8 andll limiting the separating chamber 12 at adistance from the side walls of the housing. This arrangement isparticularly simple in the utilization of an immersion frame which canbe introduced into the device from above.

Advantageously, the intermediate wall 8 forms at least partially a partof the separating chamber 12. Thus the erection of the device becomesparticularly easy.

The plate 11, which is placed parallel to the intermediate wall 8 and inthe region of the larger angle which is formed by the intermediatebottom 6 and the intermediate wall 8, and which, together with theintermediate wall, forms the separating chamber 12, is adjustable withrespect to the intermediate wall in which corresponding means, such asstuds, are provided on the intermediate wall and return-leadingelements, such as boreholes, which are suitable for this areprovided onthe plate. These have not been represented for purposes of simplicity.In this manner there is achieved in the simplest manner the variation ofthickness of the carrier substance in the separating chamber 12according to the particular requirements.

The electrodes 13 and 14 which are provided on the inside housingbackwall 3, preferably in a recess for each one, are provided withconnecting elements (not shown) which project through the backwall ofthe housing. As will be described in greater detail later on, therecesses in which the electrodes are placed are connected to the outsideair preferably by means of a canal which'runs upwards in the housingbackwall so that air bubbles rising on the electrode can escape. Theremovability of the lid 2 and the front sidewall 4 of the housing issolved by any known manner. The tightness of the removable parts withrespect to the rest of the housing is also achieved by any known means.

The described arrangement makes it possible to provide means forpressing attachments onto the lower limit of the separating chamber inthe partial space located below the separating chamber 12. These meanscan be used to close the bottom side of the separating chamber by meansof a special attachment in order to make possible the already-describedintroduction of the carrier substance inside of the device. Preferably,the pressing means are shaped in the manner ofa lifting stand 17 whichcan be easily activated from the front with the front side wall 4removed. A motorized height adjustability of the lifting stand ispossible, but it is hardly necessary in practice since the manualactivation, for example, by means of one or more threaded spindles l8and spindle nuts 19 can be carried out with few movements of the hand.

The cooling of the separating chamber 12 is particularly easy andefficient, in the case of this invention, in comparison to the alreadyknown preparative electrophoresis devices which operate basically withcylindershaped separating chambers. Because of the large side surfacesof the separating chamber which is rectangular in shape in the crosssection, provision is made for a good heat exchange and it is achievedin that no significant temperature gradient occurs in the separatingchamber. It is thus provided that inside of the separating chamber notemperature-caused differences in rate of travel occur so that theindividual'bands do not undergo any distortions. It thus also becomespossible to cleanly separate from one another even the bands which arelocated close to one another.

Within the framework of the invention another form of the intermediatewall 8 and of the separating chamber 12 is'alsopossible. Thispossibility is described in further detail based on FIG. 2.

The intermediate wall 8 is provided with a window 20, which is closed orcloseable from the front side 4 of the box-shaped housing 5 through theattachment of a plate 21 and which, together with a plate 22 whichcorresponds to the plate 11, forms the separating chamber. Between theplate 21 which closes the window 20 and the window frame (intermediatewall 8),

packing means are provided (shown by dashes in FIG. 2) so that no bufferfluid can enter from the container 9 into the remaining area of thehousing 5. Preferably the side walls of the housing 5 are not used asside limit of the separating chamber 12, but between the two plates 21and 22 which form the separating chamber there are provided laterallylimiting packing means which may be composed'of glass rods 24. The glassrods 24 are quadrangular or rectangular shaped in the cross section (seeFIG. 4) and carry compressible packing means 25 running in correspondingnuts on their two sides opposite the two plates 21 and 22. When the twoplates, which are also preferably made of glass, are pressed together,then the separating chamber 12 is practically of the same material onall of its limiting walls, so that, for example, expansion phenomena orother disturbances do not cause the carrier substance to detach from thechamber walls and creeping currents to occur which could disrupt theelectrophoresis. As already shown in FIG. 1, the plate 8 or 12 whichcloses the buffer container 9 towards the front is arranged in such away that a distance is present between its upper edge and the housinglid 2, whereas the second plate 1 1 or 22 is closer to the housing lid 2than the upper edge of the previously mentioned plate 8 or 21. This thuscauses the buffer fluid in the buffer container 9 to only be able toreach the upper end of the separating chamber 12 but not the otherbuffer container 10.

It is also possible within the scope of this invention to make the twoplates which form the separating chamber 12 of the same height and tosee to it that the buffer container 9, limited by the housing backwall 3and the plate 21 which closes the window 20, together with the upperopening of the separating chamber 12 is made tight in comparison withthe other buffer container 10 through the housing lid 2. Naturally,instead of the housing lid an intermediate lid can also be used.

It can be seen from FIG. 3 that on the window frames (intermediate wall8) there are provided threaded bolts 26 projecting towards the frontside 4 of the housing 5 and onto which there are placed nuts 27. Whenthese bolts are fastened securely the outside plate 22 is pressedagainst the packing means 24, 25 so that the plate 21 which closes thewindow 20 is also pressed against the window frame. There thus occurs atight separating chamber in the housing 5 having a thickness whichcorresponds to the thickness of theglass rods 24. The thickness of theseparating chamber can be adjusted in a simple manner by varying thethickness of the glass rods. For use in connection with this inventionit may be advantageous in analytical devices to provide for a number ofpacking means (24, 25) running parallel to the lateral packings in orderto have several separating chamber stretches available under otherwiseequal iontophoresis conditions.

With the already-mentioned lifting stand 17 it is also possible toattach to the bottom side of the separating chamber 12 a so-calledelution attachment which is described in detail below. The yields in thedevice according to this invention are considerably greater than thoseof known devices which leads to the result that a considerably greatersurface can be elutriated, whereby consequently the amounts of elutionfluid which are used can be kept small. The elution takes place from thenarrow side wall of the separating chamber 12 or the elution attachmentand on the other narrow side there occurs the removal of the elutionfluid. The yields can be increased to a still greater extent by means ofmeander-shaped path. Not only is the yield in the device according tothis invention considerably increased in comparison to that of knownelectrophoresis devices but the selectivity of bands closely followingone another is also increased. This is also a result of the designaccording to this invention since, in comparison with devices withcylindrical separating chambers, a uniform elution current can beprovided. In the case of cylindrical separating chambers the elutiondoes not take place with uniform rate of flow since the feeding andremoval of the elution current through a circular shaped or circularring shaped cross section is not possible with prior art means andconsequently irregular flushings occur which have an adverse effect uponthe selectivity of the separation. In the design example which isrepresented, the plate of the lifting stand 17 is composed of a frameinto which the elution attachment 30 shown in FIG. can be inserted. Theframe is rectangular and extends over the entire separating chambercross section. It is open from below so that the buffer fluid which ispresent in the container can reach the elution attachment from below.With the aid of the lifting stand arrangement, the attachment 30 can bepressed tightly against the lower separating chamber opening.

The elution attachment 30 is composed of a first plate 31, a secondplate 32, a third plate 33, a first semipermeable membrane 34 and asecond semiperme able membrane 35. The plates 31, 32, 33 are rectangularand are provided in the center of the plates with a slit 36 which goesthrough the thickness of the plate and the dimensions of whichcorrespond to those of the bottom side of the separating chamber. Theslit 36 need not extend over the entire length of the plate.

On the narrow sides 39, 40 of the plates 31 and 32 there are leads 41,42, 43, 44 to the slit ends 36. At 41 elution fluid is fed to theelution chamber 37 formed in the plate 31 and at 43 concentrated buffersolution is fed and removed at 44. The concentrated buffer solutionserves the purpose of greatly diminishing the electrical field intensityand in this way achieves a braking effect for the substances which areto be elutriated and results in an increase in the effective flushing.An absorption of the proteins which are to be elutriated on the membrane34 and their electrophoresis through the membrane is thus effectivelyprevented. By means of the flowing of the highly concentrated bufferfluid through the buffer chamber 38 formed in the plate 32 provision ismade to keep the concentration of the buffer fluid uniform at themembrane 34.

The plate 31 in the vicinity of its upper side above the slit 36 has anet 46 which serves for supporting the gel which is located in theseparating chamber 12. Baffle rods 45 may be provided diagonally to theflow direction in the chambers 37 and 38 interchangeably from both sidesof the slit 36 in order to force a meandershaped flow. Thus, at arelatively high rate of flow only a slight drive is necessary and theobtention of highlyconcentrated eluates is possible. The baffle rods 45also serve to keep the membranes 34 and 35 tight (see FIG. 6).

It can be seen from FIG. 5 that the plates 31 and 32 must possess athickness which corresponds at least to the strength of the connections41 to 44. Thereby there is realized a relatively large elution chamber.It is, however, worthwhile to keep the elution chamber as small aspossible so that high elution speeds can be achieved with a low elutionfluid weight rate of flow in order to provide for a good dissolution andquality of the device. According to a further development of theinvention a reduction in the elution chamber volume is achieved byproviding the connections for the feeding and removal of the elutionfluid on the ends of the separating chamber 12 which are adjacent to theelution chamber. The connections are achieved by means of boreholes(from outside of the separating chamber 12 to the elution chamberinside). In this way the plate 31 can be kept very thin and consequentlythe elution chamber volume can be kept very small so that the desiredquality improvement (better dissolving power) is achieved.

FIG. 7 shows a view of the inside of the backwall 3. It can be seen thatthe electrodes 13 and 14, which are shaped in the form of a wire here,are housed in recesses 51, 52 which are milled in the backwall 3. Therecesses are somewhat wedge-shaped so that their upper edges 53 and 54rise upwards obliquely. Gas bubbles rising to the electrodes 13, 14travel along these upper edges 53, 54 which run obliquely upwards andgather at the highest point. For diverting the existing gas, therefore,at the highest points of the wedgeshaped recesses 51, 52 a channel 55 or56 is bored upwards through the backwall 3 through which the gas canescape into the open air. From FIG. 8, which shows a cross sectionthrough the backwall 3 it can be seen that the upper edges 53 and 54 arealso designed obliquely in the direction of the outside of the backwall3. In this way the recesses 51, 52 are given a defined highest positionat which the gas collects and at which point the canals 55, 56 engageinto the recesses. For purposes of simplicity only the canal 55 of theupper recess 51 is shown in FIG. 8.

The aforementioned type of arrangement and design of the electrodes 13,14, as well as their housing into recesses 51, 52 in the backwall 3 isrecommended, for example, in disc electrophoresis.

A further advantage of this invention is that the device can also beused for the method of isoelectric focusing in which particularly smallbuffer container chambersare used. It is possible with the aid of thelifting stand 17 to press a corresponding attachment 57 onto the lowerend of the separating chamber 12'which only has a small volume andcontains an electrode 63. In this way it is possible to use one and thesame device for all knownelectrophoresis methods and for analytical andpreparative purposes only by using individual attachments.

When using the method of iso-electric focusing the electrodes should notbe far from the. separating chamber so that the potential gradient liesessentially within the separating chamber which is filled with thecarrier means. The attachment of the electrode to the lower separatingchamber opening entails, however, the danger that the gas bubbles risingfrom the electrode can reach the underside of the separating chamber anddisturb the electrophoresis.

This danger is eliminated by means of the iso-electric focusingattachment according to this invention. FIG.

9 shows sectional view through the attachment 57 which is essentiallycomposed of a square-shaped container the length of which is at leastthe length of the lower opening of the separating chamber 12. At itsupper side the container has the opening 59 which creates a connectionof the lower opening of the separating chamber 12 and the inside 60 ofthe container. By means of a closeable opening (not shown) the containeror the inside 60 of the container can be filled with buffer fluid. Theattachment 57 has an adjacent chamber 61 towering above the lowerseparating chamber limit laterally and which is partially delimited fromthe remaining part of the buffer container by means of a diaphragm 62which is essentially vertical and ending over the bottom of the buffercontainer. The electrode 63 is housed in the adjacent chamber 61 and thegas bubbles which go out from it-rise to the upper limit of the adjacentchamber 61 where they can escape through a canal 64. The opening 59 atthe upper side 58 of the attachment 57 is surrounded by packing materialwhich is arranged in such a way that it can be pressed against the lowerlimit of the separating chamber 12. FIG. 1 schematically shows how thepressing can occur with the aid of the lifting stand 17. For thispurpose, there can be used vertically standing threaded bolts located atthe housing bottom 1, therefore inside of the large buffer containerwhich is not filled during an iso-electric focusing procedure. The boltscan be moved back and forth on the threaded bolts with the aid of whichthe lifting stand 17 can move back and forth on the bolts.

In the case of iso-electric focusing a correspondingly smaller buffercontainer with electrode can be attached on the upper end of theseparating chamber 12. The advantage of the small volume is thusobtained and also the danger of the gas bubble contact with the upperend of the separating chamber 12 does not occur. The elimination of thedeveloped gas by means of a ventilation canal provided over theelectrode in its adjacent chamber according to the invention isnevertheless useful so that no disturbing excess pressure occurs in thebuffer container. The adjacent chamber can be kept very small, by theway, so that it is only insignificantly different from the volume of theiso-electric focusing attachment.

Obviously, the embodiments shown are exemplary only and a wide varietyofembodiments may be devised without departing from the spirit and scopethereof.

What is claimed is:

l. A gel-electrophoresis device comprising a vertical separatingchamber, the cross section of which is elongated and has a substantiallyrectangular shape, two buffer containers, one being in communicationwith the upper end of said separating chamber, elution meansinterconnected between the lower end of said separating chamber and theother buffer container, said elution means comprising an elongatedelution chamber, means at each of the opposed ends of said elongatedchamber for supplying and removing an elution fluid to and from saidelution chamber, an electrode in each of said buffer containers, andmeans for connecting the electrodes to a source of direct current.

2. A device as claimed in claim 1 wherein said elution means furthercomprises a second elongated chamber beneath said elongated elutionchamber for receiving aconcentrated buffer fluid.

3. A device as claimed in claim 1 wherein the lower wall of saidelongated elution chamber is composed of semipermeable membranematerial.

4. A device as claimed in claim 2 wherein said second elongated chamberhas an upper and a lower wall each composed of semipermeable membranematerial, said lower wall separating said second chamber from said otherbuffer container and said upper wall constituting the lower wall of saidelution chamber.

5. A device as claimed in claim 2 and further comprising means at eachof the opposed ends of said sec- 0nd elongated chamber for supplying andremoving the concentrated buffer fluid.

6. A device as claimed in claim 1 wherein said elution means comprises aplate having a recess therein which constitutes said elongated elutionchamber, the shape of said recess conforming with the shape of the lowerend of said separating chamber and means for tightly securing the upperside of said plate to the lower end of said separating chamber.

7. A device as claimed in claim 6 wherein said recess extends throughsaid plate and further comprising means for closing the lower side ofsaid recess.

8. A device as claimed in claim 6 wherein said elution means furtherincludes a second plate beneath said first mentioned plate and having arecess therein for receiving a concentrated buffer fluid and asemipermeable membrane firmly gripped between said first and secondplates and constituting the lower wall of said elongated elutionchamber.

9. A device as claimed in claim 8 wherein the recess of said secondmentioned plate corresponds in shape to the recess of said first plate.

10. A device as claimed in claim 8 wherein said elution means furtherincludes a third plate and a second semipermeable membrane grippedbetween said third plate and said second plate to form the lower wall ofthe recess in said second plate.

11. A device as claimed in claim 10 wherein the recess of said thirdplate corresponds in shape to the recess in said first plate.

12. A device as claimed in claim 6 wherein the means for supplying andremoving an elution fluid comprises channels leading to and from theopposed ends of the recess in said plate.

13. A device as claimed in claim 8 and further comprising channel meansin said second plate for supplying and removing the concentrated bufferfluid to and from the opposed ends of the recess in said second plate.

14. A device as claimed in claim 10 wherein said first, second and thirdplates are superposed with semipermeable membranes disposedtherebetween, said recesses being in register and conforming with theshape of the lower end of said separating chamber and further comprisingmeans for pressing the superposed plates together and for pressing theassembled plates against I the bottom of said separating chamber.

15. A device as claimed in'claim land further comprising baffle rodsdisposed within said elongated elution chamber, arranged alternately onopposite sides thereof and extending diagonally to the direction of flowof the elution fluid passing therethrough.

16. A device asclaimed in claim 1 and further comprising vertical wallsdefining said separating chamber, said walls being provided withchannels for supplying and removing the elution fluid to and from saidelongated elution chamber.

17. A device as claimed in claim 1 and further comprising adjustablemeans within the other buffer container for supporting said elutionmeans and pressing it against the lower end of said separating chamber.

18. A gel electrophoresis device comprising two laterally spacedparallel, vertical walls forming a vertical separating chambertherebetween, two buffer containers on opposed sides of said separatingchamber, one being only in communication with the upper end of saidseparating chamber and the other only in communication with the lowerend of said separating chamber, whereby each container may be easilyfilled with buffer fluids to the same level in each container, anelectrode in each of said containers and means for connecting saidelectrodes to a source of direct current.

19. A device as claimed in cliam 18 wherein each of the laterally spacedwalls form one wall of each of the buffer containers.

20. A device as claimed in claim 18, wherein one of said laterallyspaced walls extends upwardly to a higher point than the other of saidwalls.

21. A device as claimed in claim 18 and further comprising a lid memberdisposed above said separating chamber, and wherein said laterallyspaced walls extend upwardly to the same level, said lid member forminga fluid tight seal with one of said laterally spaced walls.

22. A device as claimed in claim 18 and further comprising asubstantially rectangular box-like housing having a bottom and sideelements and including an intermediate bottom element above the bottomof said housing and extending partially but not less than one half ofthe distance across said housing, the bottom of one of said laterallyspaced walls being connected to the inner end of said intermediatebottom, whereby the intermediate bottom element and said one laterallyspaced wall together with side portions of said housing form one of saidbuffer containers, the other of said buffer containers being defined bythe other of said laterally spaced walls, the bottom and side elementsof said housing, and the intermediate bottom element.

23. A device as claimed in claim 22 wherein the space between saidlaterally spaced walls is adjustable.

24. A device as claimed in claim 22 wherein a portion of said otherbuffer container is disposed beneath the lower end of said separatingchamber.

25. A device as claimed in claim 22 and further comprising a detachablelid for said housing and wherein the side element of said housing spacedfrom said intermediate bottom elcment is detachable.

26. A device as claimed in claim 22 wherein the one laterally spacedwall forming a part of said one buffer container is provided with awindow therethrough, and further comprising a cover plate for coveringsaid window on that side facing said separating chamber and means forsecuring said cover plate over said window in said one spaced wall, saidmeans also securing said other spaced wall in spaced relation to saidcover plate to form said separating chamber.

27. A device as claimed in claim 26 and further comprising packing meansbetween said cover plate and said other laterally spaced wall.

28. A device as claimed in claim 27 wherein said packing means comprisesat least two groups of vertically extending parallel members, saidgroups being spaced apart to divide said separating chamber into atleast two parallel, vertical chambers, said packing means being composedof the same material as the material facing said separating chamber.

29. A device as claimed in claim 26 and further comprising a lid memberdisposed above said separating chamber, said lid member forming a fluidtight seal with the top of said other-spaced wall to preventcommunication between said other buffer container and said one buffercontainer.

30. A device as claimed in claim 26 wherein said securing meanscomprises bolts extending outwardly from said one laterally spaced walland through said cover plate and said other laterally spaced wall andnuts for securing the. cover plate and spaced walls together.,

31. A device as claimed in claim 22 and further comprising a recess onthe inside of one wall of each of said two buffer containers forreceiving'each of said electrodes.

32. A device as claimed in claim 18 wherein a wall of the other buffercontainer in communication with the lower end of said separation chamberis provided with a recess for receiving one of said electrodes.

33. A device as claimed in claim 32 wherein the wall of the one buffercontainer in communication with the upper end of said separation chamberis provided with a recess for receiving the other of said electrodes.

34. A device as claimed in claim 31 wherein each of said recesses in thewalls is provided with an upwardly inclined upper surface and said wallsare provided with channels extending from the uppermost part of saidrecesses to the top of said walls for removing gas bubbles formed atsaid electrodes.

35. A device as claimed in claim and further comprising detachablecooling means disposed in each of said buffer containers adjacent saidlaterally spaced walls for cooling said separation chamber.

36. A device as claimedin claim 35 wherein said cooling means comprisescooling plates arranged parallel to said laterally spaced walls.

2. A device as claimed in claim 1 wherein said elution means furthercomprises a second elongated chamber beneath said elongated elutionchamber for receiving a concentrated buffer fluid.
 3. A device asclaimed in claim 1 wherein the lower wall of said elongated elutionchamber is composed of semipermeable membrane material.
 4. A device asclaimed in claim 2 wherein said second elongated chamber has an upperand a lower wall each composed of semipermeable membrane material, saidlower wall separating said second chamber from said other buffercontainer and said upper wall constituting the lower wall of saidelution chamber.
 5. A device as claimed in claim 2 and furthercomprising means at each of the opposed ends of said second elongatedchamber for supplying and removing the concentrated buffer fluid.
 6. Adevice as claimed in claim 1 wherein said elution means comprises aplate having a recess therein which constitutes said elongated elutionchamber, the shape of said recess conforming with the shape of the lowerend of said separating chamber and means for tightly securing the upperside of said plate to the lower end of said separating chamber.
 7. Adevice as claimed in claim 6 wherein said recess extends through saidplate and further comprising means for closing the lower side of saidrecess.
 8. A device as claimed in claim 6 wherein said elution meansfurther includes a second plate beneath said first mentioned plate andhaving a recess therein for receiving a concentrated buffer fluid and asemipermeable membrane firmly gripped between said first and secondplates and constituting the lower wall of said elongated elutionchamber.
 9. A device as claimed in claim 8 wherein the recess of saidsecond mentioned plate corresponds in shape to the recess of said firstplate.
 10. A device as claimed in claim 8 wherein said elution meansfurther includes a third plate and a second semipermeable membranegripped between said third plate and said second plate to form the lowerwall of the recess in said second plate.
 11. A device as claimed inclaim 10 wherein the recess of said third plate corresponds in shape tothe recess in said first plate.
 12. A device as claimed in claim 6wherein the means for supplying and removing an elution fluid compriseschannels leading to and from the opposed ends of the recess in saidplate.
 13. A device as claimed in claim 8 and further comprising channelmeans in said second plate for supplying and removing the concentratedbuffer fluid to and from the opposed ends of the recess in said secondplate.
 14. A device as claimed in claim 10 wherein said first, secondand third plates are superposed with semipermeable membranes disposedtherebetween, said recesses being in register and conforming with theshape of the lower end of said separating chamber and further comprisingmeans for pressing the superposed plates together and for pressing theassembled plates against the bottom of said separating chamber.
 15. Adevice as claimed in claim 1 and further comprising baffle rods disposedwithin said elongated elution chamber, arranged alternately on oppositesides thereof and extending diagonally to the direction of flow of theelution fluid passing therethrough.
 16. A device as claimed in claim 1and further comprising vertical walls defining said separating chamber,said walls being provided with channels for supplying and removing theelution fluid to and from said elongated elution chamber.
 17. A deviceas claimed in claim 1 and further comprising adjustable means within theother buffer container for supporting said elution means and pressing itagainst the lower end of said separating chamber.
 18. A gelelectrophoresis device comprising two laterally spaced parallel,vertical walls forming a vertical separating chamber therebetween, twobuffer containers on opposed sides of said separating chamber, one beingonly in communication with the upper end of said separating chamber andthe other only in communication with the lower end of said separatingchamber, whereby each container may be easily filled with buffer fluidsto the same level in each container, an electrode in each of saidcontainers and means for connecting said electrodes to a source ofdirect current.
 19. A device as claimed in cliam 18 wherein each of thelaterally spaced walls form one wall of each of the buffer containers.20. A device as claimed in claim 18, wherein one of said laterallyspaced walls extends upwardly to a higher point than the other of saidwalls.
 21. A device as claimed in claim 18 and further comprising a lidmember disposed above said separating chamber, and wherein saidlaterally spaced walls extend upwardly to the same level, said lidmember forming a fluid tight seal with one of said laterally spacedwalls.
 22. A device as claimed in claim 18 and further comprising asubstantially rectangular box-like housing having a bottom and sideelements and including an intermediate bottom element above the bottomof said housing and extending partially but not less than one half ofthe distance across said housing, the bottom of one of said laterallyspaced walls being connected to the inner end of said intermediatebottom, whereby the intermediate bottom element and said one laterallyspaced wall together with side portions of said housing form one of saidbuffer containers, the other of said buffer containers being defined bythe other of said laterally spaced walls, the bottom and side elementsof said housing, and the intermediate bottom element.
 23. A device asclaimed in claim 22 wherein the space between said laterally spacedwalls is adjustable.
 24. A device as claimed in claim 22 wherein aportion of said other buffer container is disposed beneath the lower endof said separating chamber.
 25. A device as claimed in claim 22 andfurther comprising a detachable lid for said housing and wherein theside element of said housing spaced from said intermediate bottomelement is detachable.
 26. A device as claimed in claim 22 wherein theone laterally spaced wall forming a part of said one buffer container isprovided with a window therethrough, and further comprising a coverplate for covering said window on that side facing said separatingchamber and means for securing said cover plate over said window in saidone spaced wall, said means also securing said other spaced wall inspaced relation to said cover plate to form said separating chamber. 27.A device as claimed in claim 26 and further comprising packing meansbetween said cover plate and said other laterally spaced wall.
 28. Adevice as claimed in claim 27 wherein said packing means comprises atleast two groups of vertically extending parallel members, said groupsbeing spaced apart to divide said separating chamber into at least twoparallel, vertical chambers, said packing means being composed of thesame material as the material facing said separating chamber.
 29. Adevice as claimed in claim 26 and further comprising a lid memberdisposed above said separating chamber, said lid member forming a fluidtight seal with the top of said other-spaced wall to preventcommunication between said other buffer container and said one buffercontainer.
 30. A device as claimed in claim 26 wherein said securingmeans comprises bolts extending outwardly from said one laterally spacedwall and through said cover plate and said other laterally spaced walland nuts for securing the cover plate and spaced walls together.
 31. Adevice as claimed in claim 22 and further comprising a recess on theinside of one wall of each of said two buffer containers for receivingeach of said electrodes.
 32. A device as claimed in claim 18 wherein awall of the other buffer container in communication with the lower endof said separation chamber is provided with a recess for receiving oneof said electrodes.
 33. A device as claimed in claim 32 wherein the wallof the one buffer container in communication with the upper end of saidseparation chamber is provided with a recess for receiving the other ofsaid electrodes.
 34. A device as claimed in claim 31 wherein each ofsaid recesses in the walls is provided with an upwardly inclined uppersurface and said walls are provided with channels extending from theuppermost part of said recesses to the top of said walls for removinggas bubbles formed at said electrodes.
 35. A device as claimed in claimand further comprising detachable cooling means disposed in each of saidbuffer containers adjacent said laterally spaced walls for cooling saidseparation chamber.
 36. A device as claimed in claim 35 wherein saidcooling means comprises cooling plates arranged parallel to saidlaterally spaced walls.